TY - JOUR
T1 - Oligomers of the Cytoplasmic Fragment from the Escherichia coli Aspartate Receptor Dissociate through an Unfolded Transition State
AU - Seeley, Stacy K.
AU - Wittrock, Gunther K.
AU - Thompson, Lynmarie K.
AU - Weiss, Robert M.
N1 - The kinetic and equilibrium properties of a clustering process were studied as a function of temperature for two point mutants of a 31 kDa fragment derived from the cytoplasmic region of the Escherichia coli aspartate receptor (C-fragment), which were shown previously to have a greater tendency to form clusters relative to the wild-type C-fragment [Long, D.
PY - 1996/12/17
Y1 - 1996/12/17
N2 - The kinetic and equilibrium properties of a clustering process were studied as a function of temperature for two point mutants of a 31 kDa fragment derived from the cytoplasmic region of the Escherichia coli aspartate receptor (C-fragment), which were shown previously to have greater tendency to form clusters relative to the wild-type C-fragment [Long, D. G., and Weis, R. M. (1992) Biochemistry 31, 9904-9911]. The clustering equilibria were different for the two C-fragment. Monomers of a serine 461 to leucine (S461L) mutant C-fragment were in equilibrium with dimers, while monomers of a S325I C-fragment were in equilibrium with trimers. The positive values for ΔH°, ΔS°, and ΔC(p)°of dissociation estimated from a van't Hoff analysis, and the differences in the CD spectral of isolated monomers and oligomers, demonstrated that the monomers were less well-folded than the clustered forms. The oligomer dissociation rate exhibited a marked temperature dependence over the range from 4 to 30 °C and was remarkably slow at low temperature; e.g. t 1/4 of dimer dissociation for the S46IL, C-fragment was 85 h at 4 °C. The values for ΔH°((+)), ΔS°((+)), and ΔC)(p)°((+)) derived from the temperature dependence of the dissociation rate were comparable to the corresponding parameters determined in a DSC study of C-fragment denaturation [Wu, J., Long, D. G. and Weis, R. M. (1995) Biochemistry 34, 3056-3065], which indicated that the transition state resembled thermally denatured C-fragment. Octyl glucoside accelerated the dissociation rate by 3-5 fold presumably by lowering the barrier to dissociation. The acceleration and the positive value of ΔC(p)°((+)) were interpreted as evidence for an increase in solvent accessible hydrophobic groups in the transition date. The molecular basis for the slow rate of dissociation is proposed to result from the conversion of intermolecular coiled coils in the oligomers to an intramolecular coiled coil in the monomer.
AB - The kinetic and equilibrium properties of a clustering process were studied as a function of temperature for two point mutants of a 31 kDa fragment derived from the cytoplasmic region of the Escherichia coli aspartate receptor (C-fragment), which were shown previously to have greater tendency to form clusters relative to the wild-type C-fragment [Long, D. G., and Weis, R. M. (1992) Biochemistry 31, 9904-9911]. The clustering equilibria were different for the two C-fragment. Monomers of a serine 461 to leucine (S461L) mutant C-fragment were in equilibrium with dimers, while monomers of a S325I C-fragment were in equilibrium with trimers. The positive values for ΔH°, ΔS°, and ΔC(p)°of dissociation estimated from a van't Hoff analysis, and the differences in the CD spectral of isolated monomers and oligomers, demonstrated that the monomers were less well-folded than the clustered forms. The oligomer dissociation rate exhibited a marked temperature dependence over the range from 4 to 30 °C and was remarkably slow at low temperature; e.g. t 1/4 of dimer dissociation for the S46IL, C-fragment was 85 h at 4 °C. The values for ΔH°((+)), ΔS°((+)), and ΔC)(p)°((+)) derived from the temperature dependence of the dissociation rate were comparable to the corresponding parameters determined in a DSC study of C-fragment denaturation [Wu, J., Long, D. G. and Weis, R. M. (1995) Biochemistry 34, 3056-3065], which indicated that the transition state resembled thermally denatured C-fragment. Octyl glucoside accelerated the dissociation rate by 3-5 fold presumably by lowering the barrier to dissociation. The acceleration and the positive value of ΔC(p)°((+)) were interpreted as evidence for an increase in solvent accessible hydrophobic groups in the transition date. The molecular basis for the slow rate of dissociation is proposed to result from the conversion of intermolecular coiled coils in the oligomers to an intramolecular coiled coil in the monomer.
UR - https://pubs.acs.org/doi/10.1021/bi961749i
U2 - 10.1021/bi961749i
DO - 10.1021/bi961749i
M3 - Article
VL - 35
JO - Biochemistry
JF - Biochemistry
ER -